Refined Method for Droplet Microfluidics-Enabled Detection of Plasmodium falciparum Encoded Topoisomerase I in Blood from Malaria Patients

Micromachines

Marianne Smedegaard Hede⁵  Patricia Nkem Okorie⁶  Signe Kirk Fruekilde¹  Søren Fjelstrup¹  Jonas Thomsen⁵  Oskar Franch¹  Cinzia Tesauro¹  Magnus Tobias Bugge¹  Mette Christiansen⁷  Stéphane Picot⁸  Felix Lötsch⁴  Ghyslain Mombo-Ngoma⁴  Johannes Mischlinger⁴  Ayôla A. Adegnika⁴  Finn Skou Pedersen¹  Yi-Ping Ho¹  Eskild Petersen²  Magnus Stougaard⁵  Michael Ramharter⁴  Birgitta Ruth Knudsen¹  Andrew deMello³ 

[1] Department of Molecular Biology and Genetics, University of Aarhus, 8000 Aarhus C, Denmark;Department of Infectious Diseases, Aarhus University Hospital, 8200 Aarhus N, Denmark;;Zymonostics, 8000 Aarhus C, DenmarkCentre de Recherches Médicales de Lambaréné, Lambaréné, Gabon;Zymonostics, 8000 Aarhus C, Denmark;Institute for Advanced Medical Research and Training, College of Medicine, University of Ibadan, 200001 Ibadan, Nigeria;Department of Clinical Immunology, Aarhus University Hospital, 8200 Aarhus N, Denmark;Malaria Research Unit, SMITh, ICBMS, UMR 5246 CNRS-INSA-CPE-UCBL1 & Institut de Parasitologie et Mycologie Médicale, Hospices Civils de Lyon, 69373 Lyon Cedex 08, France;

关键词: malaria;    point-of-care;    droplet microfluidics;    diagnostics;    topoisomerase;   

DOI  :  10.3390/mi6101432

来源: mdpi

【 摘 要 】

Rapid and reliable diagnosis is essential in the fight against malaria, which remains one of the most deadly infectious diseases in the world. In the present study we take advantage of a droplet microfluidics platform combined with a novel and user-friendly biosensor for revealing the main malaria-causing agent, the Plasmodium falciparum (P. falciparum) parasite. Detection of the parasite is achieved through detection of the activity of a parasite-produced DNA-modifying enzyme, topoisomerase I (pfTopoI), in the blood from malaria patients. The assay presented has three steps: (1) droplet microfluidics-enabled extraction of active pfTopoI from a patient blood sample; (2) pfTopoI-mediated modification of a specialized DNA biosensor; (3) readout. The setup is quantitative and specific for the detection of Plasmodium topoisomerase I. The procedure is a considerable improvement of the previously published Rolling Circle Enhanced Enzyme Activity Detection (REEAD) due to the advantages of involving no signal amplification steps combined with a user-friendly readout. In combination these alterations represent an important step towards exploiting enzyme activity detection in point-of-care diagnostics of malaria.

【 授权许可】

CC BY   
© 2015 by the authors; licensee MDPI, Basel, Switzerland.